1. The Field of the Invention
This invention relates generally to methods for testing reactions of substances on human skin. In particular, the present invention relates to such testing methods which are capable of using living human skin to test the properties of various substances and medicaments without directly involving a human subject.
2. Description of the Art
The behavior of human skin in various environments and its reaction to chemicals of all types is of particular interest in medicine and other related areas of technology. This is true because of the importance of the skin in performing a wide range of functions critical to the maintenance and well being of the human organism as a whole.
The primary functions of skin are to provide the body with protection and to maintain a constant environment for the organism. The skin protects the body against the influence of mechanical, thermal, chemical, and electrochemical agents encountered in the environment, and it also protects the body against the invasion of microorganisms.
The skin regulates body temperature by sweat secretion and by varying the circulation of the blood through the skin. The skin acts as a housing for the neurosensory organs critical to the senses of touch, cold, heat, and pain. The skin provides a site for the synthesis of various substances, including vitamin D, and also functions to prevent the body from losing excessive amounts of fluids, heat, and the like. The critical importance of the skin can be readily appreciated by the numerous important functions performed by the skin.
Because of the importance of the skin in protecting an individual from the environment and in performing other critical functions, the study of the skin in an important area of medical science. The study of the skin clearly has the potential of providing additional insights into the mechanism of disease and the nature of the reaction between harmful agents and the body. An understanding of the interaction of skin with various toxic agents, including carcinogens, is important to the understanding of disease functions and methods for preventing disease. Study of the interaction of the skin and the environment also provides insight into the characteristics of a safe and healthy environment.
In understanding the function of the skin and the reaction of substances in the environment with the skin, it is apparent that the study of the penetration of the skin by various substances is important. It is generally found, as would be expected, that smaller molecules can penetrate the skin faster than larger ones.
Furthermore, non-ionized molecules are more effective in penetrating the skin than are ionized compounds. Finally, it has been found that very large molecules such as proteins and polysaccharides penetrate the skin very slowly. However, beyond the generalities mentioned above, it would be desirable to perform actual tests on the skin in situ in order to come to a full understanding as to the action of any particular molecule on the skin.
Several different approaches have been developed for testing the effects of substances in contact with the skin. One such method involves simply placing a quantity of the substance to be tested on the skin of a human subject. At a predetermined time later, a sample of body fluid, such as urine or blood, is extracted and analyzed. From the resulting analysis, some understanding can be gained of the ability of the substance to penetrate the skin.
Several disadvantages are apparent when employing such a method of testing. One obvious problem is that when testing potentially harmful agents, including new pharmaceuticals, toxic substances, or carcinogens, it is not ethical or proper to use a living human subject. The danger of harm to the human subject is simply too great.
Even if problems generally related to safety could be ignored, testing using human subject still has serious difficulties. One primary difficulty is that the accuracy of the test is extremely limited. For example, the substance to be tested must penetrate the skin and then enter the blood stream. Once the substance enters the blood stream it will pass through the kidneys, the liver, and other internal organs. These organs will tend to sweep the impurity out of the blood stream. In addition, it is found that various mechanisms within the skin itself may remove the substance, so that it is difficult to obtain an accurate indication of the actual absorption of the substance by the skin.
In order to overcome the problem of administering potentially harmful substances to human subjects, tests on animals have often been substituted. As would be expected, in addition to the problems discussed above, difficulties are encountered in animal tests because of the extreme differences between animal skin and human skin. Some of the differences include the differences in thickness of the skin, differences hair density, and differences in the number of sebaceous and eccrine glands.
It has been found that the skin of the domestic pig most closely approaches human skin. However, even pig skin is substantially different from human skin, and test results obtained on pigs cannot necessarily be directly correlated to humans.
Because of the problems described above in testing substances on live human subjects and on animal subjects, researchers have developed other test methods. One widely used testing method simply involves a sample of dead skin removed from a human subject. The skin is placed in a test apparatus where the desired test substance is placed on the skin and its penetration through the dead skin sample is measured. Essentially a controlled amount of the substance is placed on top of the sample and the presence of the substance below the sample is subsequently measured.
In order to use the results of this test method, it is generally necessary to make the assumption that dead skin behaves in the same manner as living skin and that the effects of circulation through the skin are negligible. This assumption, however, is manifestly incorrect. Living skin dynamically removes substances which enter through the skin surface. Thus, the effect of blood flow through the living skin, as well as many other factors, make test results using dead skin samples unpredictable when correlated to living subjects.
Thus, the art is presently lacking an adequate system for testing the reaction of substances on the skin. In particular, there exists no satisfactory method for testing the action of substances on living human skin. As discussed above, animal experiments and experiments with dead human skin do not provide satisfactory results.
The art is lacking testing methods which facilitate the study of the metabolic capacity of the skin, the interaction of the skin with toxic substances, and the study of the skin as a barrier to the delivery of pharmaceuticals. Such testing methods could provide an understanding of the reaction of the skin with various drugs in order to study the potential for systemic drug delivery through the skin. For example, such testing methods could provide insight into the administration of medicaments using iontophoresis and other similar noninvasive administration techniques.
An adequate testing method would provide a method for studying the metabolism of hormones, cytokins, or other substances in the skin and could provide additional insight into the inflammatory processes. In addition, an adequate testing method using the microcirculation of the skin would be helpful in understanding the distribution of various agents through the microcirculatory system. This would aid in defining the role of microcirculation through the skin in overall physiological processes.
Thus, what is needed in the art are improved methods and experimental models and testing the reaction of substances on the human skin. In particular, what is needed in the art are methods and testing models for testing the reaction of substances with living human skin. It would be a further advancement in the art to provide such methods and apparatus which would allow a researcher to directly access the blood supply to a skin sample. Specifically, it would be an advancement in the art to provide such testing methods and models which would provide for the testing of blood directly below the skin, without allowing the blood to first travel through the organism as a whole.
Such methods and testing models are disclosed and claimed herein.